ABSTRACT
Previous evidence suggests that the thromboembolic risk is greater among patients with COVID-19 than among those affected by other types of acute respiratory distress syndrome (ARDS). However, such comparison has been primarily based on historical cohorts. In order to reduce the possible influence of such selection bias, the main goal of this study was to evaluate thromboembolic events in patients with COVID-19 and other ARDS hospitalized in the same time period. For this reason, we have selected patients admitted from March to June, 2020 at the UNICAMP Clinical Hospital who met the ARDS clinical criteria established by the Brazilian Ministry of Health and the Berlin Definition by presenting two or more flu-like symptoms and at least one ARDS-specific manifestation (dyspnea, persistent chest pressure, oxygen saturation lower than 95% at hospital admission, or lip/face cyanosis). Symptom onset or worsening occurred 30 days before hospital admission at the latest, and COVID-19 diagnosis was confirmed or excluded by at least 2 real time polymerase chain reactions or enzyme-linked immunosorbent assays. Descriptive analysis, chi-square and t-tests, as well as binary logistic regression, were used to compare COVID-19 and non-COVID-19 patients. Of the 253 patients hospitalized due to ARDS during this period, 101 COVID-19 and 102 non-COVID-19 patients were included in this study. The remaining patients were excluded due to incomplete medical records (n=16) or absence of COVID-19 testing results (n=34). Table 1 demonstrates the included patients' demographic and clinical baseline features. Both COVID-19 and non-COVID-19 groups showed similar baseline risk of hospital-associated thrombosis (assessed by reduced mobility within the past 3 days or more, previous thromboembolism event, recognized “thrombophilia”, and infarction, stroke, trauma or surgery within the past 4 weeks) and oxygen saturation at admission (COVID-19: 92% IQR 90% to 96%;non-COVID-19: 94% IQR 91% to 97%, P=0.44). However, the need for invasive oxygenation support (37.6% vs. 14.7%, P=0.0002) and vasoactive drugs (44.6% vs. 21.6%, P=0.0006) was greater in COVID-19 than in non-COVID-19 patients. Accordingly, those infected by SARS-CoV-2 were more frequently admitted in ICU (55.4% vs. 40.2%, P=0.04) and for a longer period of time (13 days IQR 6 to 22 vs. 3 days IQR 2 to 8.3, P=0.02) than those affected by other types of ARDS. In comparison to the non-COVID-19 group, the COVID-19 group's median total hospital stay was more lasting (15 days IQR 6 to 30.5 vs. 7 days IQR 3 to 16.3, P<0.0001), and its death rate, higher (27.7% vs. 14.7%, P=0.03), as shown in Table 2. With respect to coagulation markers (Table 3), activated partial thromboplastin time and C-reactive protein levels were greater in COVID-19 than in non-COVID-19 patients, while the latter presented higher median platelet counts. There was no statistically significant difference between both study groups in regards to prothrombin time, fibrinogen, and D-dimer levels (COVID-19: 1488 ng/mL IQR 726.5 to 3476;non-COVID-19: 1773 ng/mL IQR 807.5 to 4153.8, P=0.57). Although thromboprophylaxis was more commonly administered to COVID-19 (76.2%) than non-COVID-19 patients (41.2%, P<0.0001), the incidence of thromboembolic events confirmed by imaging examination was similar between groups even after adjusting for multiple factors (age, sex, thromboprophylaxis use, arterial hypertension, and cancer): there were 7 confirmed events in 7 non-COVID-19 patients, and 13 confirmed events in 9 COVID-19 patients (adjusted OR 0.74, 95% CI 0.24-2.25, P=0.59). Table 4 demonstrates the characteristics of such thrombotic manifestations. By analyzing patients hospitalized in the same time period, we have found that although high, the thromboembolic risk in COVID-19 is similar to that in other types of ARDS, indicating that a hypercoagulable state is inherent to ARDS in general. Additionally, the obtained results show that the use of thromboprophylaxis was significantly higher among COVID-19 patients, and that there was no tatistically relevant difference between COVID-19 and non-COVID-19 patients' D-dimer levels, a commonly used coagulation marker. Such findings provide a better understanding of the thromboembolic risk associated with SARS-CoV-2 infection, and suggest that previous evidence of higher thrombosis rates in COVID-19 suffered bias from the use of historical cohorts. [Formula presented] Disclosures: No relevant conflicts of interest to declare.
ABSTRACT
Background: the pathogenesis of severe COVID-19 involves the deregulated activation of different compartments of immunothrombosis, which are otherwise important for pathogen eradication and tissue repair. Coagulation activation, angiogenesis and alterations of endothelial barrier (EB) are elements of immunothrombosis that have been shown to be involved in the pathogenesis of COVID-19. Angiopoietins (Ang) 1 and 2 and their receptor Tie2 and VEGF-A are well-known pro-angiogenic mediators that, during inflammation also mediate EB disruption. Recently, it has also been demonstrated that the Ang/Tie2 pathway is involved in coagulation activation. Here we explored whether increased levels of angiogenesis/EB regulators (which have been previously associated with disease severity in COVID-19) are also associated with both EB disruption and coagulation activation in this condition. Methods: the study population consisted of 30 patients with COVID-19 confirmed by RT-PCR and presenting typical CT findings admitted due to hypoxemia. Thirty sex- and age-matched healthy individuals were recruited at the same time, from the same geographic region. Patients were part of a clinical trial (REBEC: U1111-1250-1843) but samples were obtained before any study intervention, within 24 hours from diagnosis confirmation. Circulating levels of angiogenesis/EB regulation mediators and coagulation biomarkers were measured by commercial assays (immunological or functional). Monolayers of endothelial cells from umbilical veins (HUVECs) or lung (HULECs) were used for measurement of EB integrity using an impedance sensor system (ECIS, Electric Cell-substrate Impedance Sensing System). Cells were stimulated with serum from patients or healthy individuals and EB integrity was continuously monitored for 36 hours. Clinical outcomes were obtained from the digital medical records. Results: mean length of hospital stay (LOS) was 12.9 ± 9.8 days. Twelve patients (40%) required intensive care (ICU) and 28/30 patients survived. Mean D-dimer was 3,609 ± 14,440 ng/mL. Circulating levels of Ang1, Ang2, sTie2 and VEGF-A were all significantly increased in patients compared to healthy individuals (Ang1: 463.2 ± 194.6 vs 237.4 ± 104.9 pg/mL, p<0.0001;Ang2: 1,926 (1,275 - 3,134) vs 1,215 (9 - 1,440 pg/mL), p<0.0001;Tie2: 10,753 ± 2,377 vs 8,603 ± 1,851 pg/mL, p<0.0001 and VEGF-A: 94.7 (73.4 - 116.0) vs 45.9 (39.7 - 57.0 pg/mL), p<0.0001.). In contrast, soluble VE-cadherin levels were decreased in patients compared to healthy individuals (1,234 ± 318 vs 1,539 ± 363 ng/mL, P=0.001). Serum from COVID-19 patients induced decreases of EB integrity in monolayers of both HUVECs and HULECs as early as 15 minutes, lasting up to 5 hours after stimulation (figure 1). The magnitude of EB disruption was correlated with clinically relevant outcomes such as time of ICU stay and LOS (figure 1). Interestingly, levels of Ang1, Ang2 and soluble VE-cadherin levels were also significantly correlated with the magnitude of EB disruption, as well as with biomarkers of coagulation activation such as fibrinogen, Von Willebrand Factor antigen levels, PAI-1, P-selectin and urokinase receptor (uPAR). Conclusions: Ang-1/Ang-2 mediated Tie2 signaling has been shown to be important for the fine regulation of barrier integrity and coagulation activation at the endothelial level, which are two critical elements of immunothrombosis. Our results provide evidence supporting that the interplay between these processes can play a role in the mechanisms driving COVID-19 severity, and suggest that targeting the Ang/Tie2 and VEGF-A pathways could be attractive strategies to modulate not only changes of the alveolar-capillary barrier, but also of coagulation activation in COVID-19. Figure 1. In (a), endothelial barrier (EB) integrity of HUVEC monolayers upon stimulation by serum from COVID-19 patients and healthy individuals (n=27-30 per group). The lower the normalized resistance, the higher the magnitude of EB disruption. Significant differences (* to ****) are evident from 15 min to 5 hours (An va corrected for multiple comparisons). In the lower panels, the correlation of EB disruption with clinically relevant outcomes such as length of hospital stay (b) and days of intensive care (c) are shown. Negative correlations (Spearman test) indicate that the magnitude of EB disruption is associated with worse outcomes. [Formula presented] Disclosures: No relevant conflicts of interest to declare.
ABSTRACT
Background: The high risk of venous thromboembolism (VTE) is a hallmark of COVID-19, particularly in intensive care units (ICU) patients. However, the magnitude of this risk is a matter of debate due to studies heterogeneity, significant changes on VTE management in COVID-19 era and scarce evidence of VTE risk in ICU patients with pneumonia in the pre-COVID-19 era. Aims: To evaluate the VTE risk in the pre-COVID-19 era in a large ICU database. Methods: Data of consecutive pneumonia patients admitted to ICU were retrieved from the Medical Information Mart for Intensive Care III (MIMIC-III) . VTE incidence during ICU stay was described. The association of thromboprophylaxis and VTE risk was determined by logistic regression, adjusted for age, sex, SOFA score, pneumonia diagnosis and type of ICU. Results: Among 6,842 pneumonia patients admitted to ICU, the median ICU stay was 11 (IQR 6-20) days. Tables 1 and 2 summarizes patients' characteristics and outcomes. 486 patients were diagnosed with VTE after a median of 3 (IQR 1-11) days in ICU. The overall cumulative incidence of VTE was 7% (95%CI 6.4-7.6), corresponding to a daily VTE incidence of 0.51% (95%CI 0.47-0.56). 1788 patients received thromboprophylaxis (out of 2958 for whom that data was available). The cumulative incidence of VTE was 10.7% (95%CI 8.9-12.6) among patients without thromboprophylaxis and 6.5% (95%CI 5.4-7.8) among those with thromboprophylaxis. Overall mortality was 19.3%, that was similar among patients with and without VTE (20.6% and 19.2%, respectively). (Table Presented) Conclusions: In pre-COVID-19 era, VTE rates in ICU patients with pneumonia was not substantially different from those reported in COVID-19 when VTE diagnosis is based on clinical suspicion. The risk of VTE was reduced by 46% with thromboprophylaxis. These findings can serve as comparator for future studies aiming at evaluating the impact of VTE on COVID-19.
ABSTRACT
Background: Hemostasis activation is a hallmark of COVID-19, and an integral part of a broader host response known as immunothrombosis. Microvesicles (MVs) are lipid-bilayer particles released from cells, and thought to be relevant mediators of immunothrombosis, due to their capacity to transfer proteins such as tissue factor (TF) and fibrinolytic mediators between different cells. Aims: To quantify platelet, endothelial cell (EC) and red blood cell (RBC) MVs in plasma from patients with COVID-19 and to explore their association with immunothrombosis mediators. Methods: Samples were obtained from patients admitted to a COVID-19 ward as part of a clinical trial. Samples were collected at admission, before any study intervention. MVs levels were measured in double centrifuged platelet poor plasma by flow cytometry. Coagulation, fibrinolysis and endothelial activation markers were measured by functional or immunological methods. The study was approved by the IRB and all participants provided written informed consent. Results: Data were obtained from 30 patients and 30 age and sexmatched healthy individuals. Mean time from symptoms onset and length of hospital stay (LOS) were 8.1 ± 2.3 days and 12.9 ± 9.8 days respectively. Twelve patients (40%) required intensive care (ICU), and 28/30 patients survived. Mean D-dimer was 3,609 ± 14,440 ng/mL. Platelet, EC and RBC MV counts are shown in Table 1. No association was observed between MV counts and clinical markers of disease severity such as LOS (total or in ICU), oximetry, and lung CT extension score. Associations between MV counts and biomarkers of coagulation, fibrinolysis and EC activation are shown in Table 2. Conclusions: Increased levels of MV from platelets and EC were observed in COVID-19. The association of these MVs with markers of coagulation and fibrinolytic activation, and with elements of the Tie2/Ang1 pathway warrant additional studies on the contribution of these pathways to the pathogenesis of COVID-19.
ABSTRACT
Background : Endothelial barrier (EB) disruption is an important part of immunothrombosis, allowing the access of leukocytes to inflamed tissues. Pathways involving angiopoietin (Ang) 1 and 2 and their receptor Tie2, and VEGF-A/VE-cadherin (VEC) are key regulators of EB integrity. While the association of these mediators with sepsis severity have been known for more than 15 years, it was only recently that their role in coagulation activation was described. Moreover, these proteins also mediate angiogenesis, which has been shown to be upregulated in COVID-19. Aims : To measure circulating levels of key mediators of Ang/Tie2 and VEGF-A pathways in COVID-19 patients, and to explore their association with disease severity and hemostatic activation. Methods : Samples were obtained from patients admitted to a COVID-19 ward, within 24 h from COVID-19 confirmation. EB mediator levels were measured by immunological methods (Elisa or multiplex assays). The study was approved by the IRB and all participants provided written informed consent. Results : Data were obtained from 30 patients and 30 age and sexmatched healthy individuals. Mean length of hospital stay (LOS) was 12.9 ± 9.8 days respectively, twelve patients (40%) required intensive care (ICU), and 28/30 patients survived. Mean D-dimer was 3,609 ± 14,440 ng/mL. Levels of EB mediators are shown in Table 1. Associations between these parameters with relevant clinical and laboratory markers of disease severity are shown in Table 2. Conclusions : All mediators of EB disruption were significantly elevated in COVID-19 patients. In addition, these mediators were consistently associated with proteins involved in immunothrombosis, in particular with fibrinogen, VWF:Ag, uPAR, PAI-1 and P-selectin. Clinically significant associations were observed between Ang-2 and VEGF-A and extension of lung disease (for both) and ICU stay (for VEGF-A). Additional studies are warranted to explore the crosstalk between Ang/Tie2 and VEGF-A pathways with hemostasis in COVID-19.
ABSTRACT
Background : Hemostasis activation is considered a key pathogenic element of COVID-19, as evidenced by the frequency of lung microvascular thrombosis and of venous thromboembolism. As shown in other conditions, pathways that mediate hemostasis activation during inflammation are not necessarily the same as those that drive hemostasis in non-inflammatory states. In particular, contact system and intrinsic pathway activation have been increasingly explored as potential mediators of hemostasis activation and prothrombotic states observed in both sterile and infectious inflammatory conditions. Aims : To assess the activation of contact system and intrinsic pathway activation in COVID-19. Methods : Protease:serpin complexes were measured in plasma from inpatients with COVID-19 and healthy individuals (HI). All samples were collected within 24 h of COVID-19 diagnosis. Associations with laboratory and clinical markers of hemostasis activation and disease severity were explored. The study was approved by the IRB and all participants provided written informed consent. Results : In total, 30 patients with COVID-19 and 30 age and sex-matched HI were enrolled. Main characteristics of the study population are shown in Table 1. As expected, higher levels of classical coagulation activation parameters were observed in COVID-19 patients. Contact system and intrinsic pathway activation in COVID-19 was demonstrated by increased levels of several protease:serpin complexes (Table 2). Interestingly, a consistent and significant association was observed between FIXa:antithrombin complexes with both clinical endpoints (need of ICU admission, length of ICU stay, total length of stay and extent of lung CT alterations), and with laboratory markers of immunothrombosis (neutrophil:lymphocyte ratio, C-reactive protein, D-dimer, PAP, VWF:Ag), as well as with several other activation markers of contact system and intrinsic pathway Conclusions : Contact system and intrinsic pathway activation contribute to hemostasis activation in COVID-19. The association of FIXa:antithrombin complexes with disease severity suggests that these pathways contribute to the pathogenesis of the prothrombotic state of COVID-19.